Structure and Regulation of hsp90 Chaperones

hsp90 分子伴侣的结构和调控

基本信息

批准号：
8461076
负责人：
DANIEL T GEWIRTH
金额：
$ 30.47万
依托单位：
HAUPTMAN-WOODWARD MEDICAL RESEARCH INST
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-05-01 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8461076
关键词：
ATP phosphohydrolase Affinity Bad protein Binding Biochemical Biological Biological Assay Cell secretion Cell surface Cells Cessation of life Client Complex Coupled Cyclic GMP-Dependent Protein Kinases DNA Sequence Rearrangement Development Discrimination Disease Drug Design Exhibits Family member GRP94 Goals Grant Heat-Shock Proteins 90 Lectin Length Ligand Binding Ligands Malignant Neoplasms Maps Mediator of activation protein Molecular Chaperones Molecular Conformation N-terminal Neoplastic Cell Transformation Neurofibrillary Tangles Nucleotides Oncogenic Play Positioning Attribute Property Proteins Regulation Research Role Shapes Signal Transduction Staging Structure System Testing Therapeutic Time Treatment Efficacy Weather Work X-Ray Crystallography base cancer cell cancer therapy cell killing in vivo inhibitor/antagonist insight neoplastic cell next generation novel osteosarcoma paralogous gene protein protein interaction public health relevance response small molecule steroid hormone receptor success time use transcription factor

项目摘要

DESCRIPTION (provided by applicant): The hsp90 chaperones participate in the late stage maturation of proteins involved in diverse cellular activities ranging from signaling to bacterial recognition. Cytoplasmic Hsp90 guides the maturation of steroid hormone receptors, proto-oncogenic kinases, G-proteins, and other key mediators of neoplastic transformation. Inhibitors of Hsp90 that disrupt this maturation display potent anti-cancer activity. GRP94, the ER hsp90 paralog, chaperones proteins destined for transport to the cell surface and secretion, plays a key role in the export of toxic malfolded proteins from the ER, and when targeted leads to the death of tumor cells. The activity of hsp90 chaperones is regulated by the binding of ATP and other ligands to the N-terminal domain, and the response of this domain is central to the hsp90 chaperone cycle. Yet despite their high degree of sequence and structural homology, the cytoplasmic and ER hsp90 paralogs exhibit fundamental differences in their response to regulatory ligands. Ligand dependent conformational changes in the N-terminal domain lid of cytoplasmic Hsp90 have been difficult to demonstrate, and a plausible trigger for any changes has not been identified. In GRP94, on the other hand, a conserved insertion in the lid primes this region to undergo conformational rearrangements that differ dramatically depending on the identity of the incoming ligand. These rearrangements alter the quaternary structure of the chaperone. The research in this proposal will identify the ligand dependent conformational states available to GRP94 using X-ray crystallography to visualize the structures of ligand-driven GRP94 complexes. In vivo assays will be used to assess the biological importance of these structural rearrangements. The sensitive response of the GRP94 lid to incoming ligands allows GRP94 to bind inhibitors that cannot be accommodated in cytoplasmic Hsp90 and are therefore selective for GRP94. Because selective and non-selective ligands are closely related, we will use X-ray crystallography to visualize the both classes of ligands bound to GRP94 and Hsp90 and infer their mechanism of selectivity. Because each of the hsp90 paralogs is responsible for chaperoning distinct sets of client proteins, specific targeting of one hsp90 paralog with selective inhibitors may result in higher efficacy and therapeutic control. Finally, new developments offer the prospect of characterizing novel client and co-chaperone interactions with both Hsp90 and GRP94: minimal client protein loading systems open the way to isolating client-Hsp90 complexes for biochemical and biophysical study, and the recent identification of the first GRP94 accessory factor, os9, has, for the first time, opened the door to characterizing how protein-protein interactions occur in the ER paralog.

描述（由申请人提供）：hsp90 伴侣参与蛋白质的后期成熟，这些蛋白质涉及从信号传导到细菌识别的多种细胞活动。细胞质 Hsp90 引导类固醇激素受体、原癌激酶、G 蛋白和其他肿瘤转化关键介质的成熟。破坏这种成熟的 Hsp90 抑制剂显示出有效的抗癌活性。 GRP94 是 ER hsp90 旁系同源物，是转运至细胞表面并分泌的伴侣蛋白，在从 ER 输出有毒错误折叠蛋白的过程中发挥着关键作用，当靶向时会导致肿瘤细胞死亡。 hsp90 分子伴侣的活性通过 ATP 和其他配体与 N 末端结构域的结合来调节，该结构域的响应是 hsp90 分子伴侣循环的核心。然而，尽管细胞质和 ER hsp90 旁系同源物具有高度的序列和结构同源性，但它们对调节配体的反应表现出根本差异。细胞质 Hsp90 N 端结构域盖的配体依赖性构象变化很难证明，并且尚未确定任何变化的合理触发因素。另一方面，在 GRP94 中，盖子中的保守插入促使该区域发生构象重排，构象重排根据传入配体的身份而显着不同。这些重排改变了伴侣的四级结构。该提案中的研究将使用 X 射线晶体学来识别 GRP94 可用的配体依赖性构象状态，以可视化配体驱动的 GRP94 复合物的结构。体内测定将用于评估这些结构重排的生物学重要性。 GRP94 盖对传入配体的敏感反应使 GRP94 能够结合不能容纳在细胞质 Hsp90 中的抑制剂，因此对 GRP94 具有选择性。由于选择性和非选择性配体密切相关，我们将使用 X 射线晶体学来观察与 GRP94 和 Hsp90 结合的两类配体，并推断它们的选择性机制。由于每个 hsp90 旁系同源物负责陪伴不同组的客户蛋白，因此用选择性抑制剂特异性靶向一个 hsp90 旁系同源物可能会产生更高的功效和治疗控制。最后，新的发展为表征新客户和共伴侣与 Hsp90 和 GRP94 的相互作用提供了前景：最小客户蛋白负载系统为分离客户-Hsp90 复合物以进行生化和生物物理研究开辟了道路，并且最近鉴定了第一个 GRP94辅助因子 os9 首次为描述 ER 旁系同源物中蛋白质-蛋白质相互作用的发生方式打开了大门。